Summary In-spite of effective strategies for lowering cholesterol, atherosclerosis and associated cardiovascular diseases remain a major global health burden. Work in recent years has highlighted the exciting therapeutic potential for interventions that reduce inflammation, as well as the need for identifying novel and selective targets. Our long term goal is to understand how lipid metabolism and inflammation interact, and how these interactions can be useful therapeutically. The objective of the current research proposal is to determine the role of the transcription factor Nuclear factor erythroid 2 related factor-1 (Nrf1; also known as Nfe2L1) in linking these two processes. The central hypothesis is that Nrf1 in the macrophage is a critical factor for preventing chronic inflammation and maintaining cholesterol homeostasis, and that enhancing these actions of Nrf1 may be beneficial in atherosclerosis. This is based on compelling new preliminary data showing that Nrf1 is activated in macrophages in response to inflammatory stimuli and is necessary for the expression of lipid metabolism pathway genes that are crucial for proper resolution of the inflammatory state. Previous results also showed that deletion of Nrf1 drastically impairs cellular cholesterol homeostasis.The rationale for the proposed research is that understanding the function and mechanism of macrophage Nrf1 may provide important insight into regulation of macrophage inflammation and resolution in general and reveal a novel therapeutic target for the treatment of atherosclerosis. The central hypothesis will be tested by pursuing three specific aims: 1) Examine the function of Nrf1 in macrophages; 2) Elucidate the molecular mechanism of Nrf1 action in macrophages; and 3) Explore the importance of macrophage-Nrf1 in atherosclerosis in-vivo. Under the first aim, tissue-specific inducible deletion models established and tested in the applicant's lab will be used for a thorough characterization of the function of Nrf1 in inflammatory output, lipid metabolism and resolution of inflammation. This aim will also explore the impact of cholesterol on Nrf1 function, and the reciprocal role of Nrf1 in regulating cholesterol homeostasis in macrophages. In the second aim, immunoprecipitation of readily available tagged protein coupled to DNA sequencing and MS/MS analysis will identify, protein interactors and post translational modifications of Nrf1 potentially mediating its function. The third aim will test the impact of Nrf1 deficiency and overexpression on atherosclerosis in-vivo using already established Western-diet fed Ldlr-deficient mice with either macrophage specific deletion of Nrf1, or adeno-associated-virus (AAV) mediated macrophage specific overexpression of Nrf1. The approach is innovative because it attempts to overcome issues of pro-resolving mediator delivery by using naturally existing systems within macrophages. The proposed research is significant, because current approaches to activate endogenous resolution of inflammation in the context of atherosclerosis are limited, and this study might provide important insight into the role of the resolution in atherosclerosis and reveal a novel therapeutic target.